The overall goal of this proposal is to explore the molecular and electrophysiological role of NFATC1 as a novel atrial fibrillation (AF) susceptibility gene and to define the previously unknown contribution of NFATC1 to atrial excitability. AF, the most common sustained arrhythmia encountered in clinical practice, has an economic burden exceeding $7 billion in annual health care costs. Emerging evidence implicates cardiac transcription factors in the pathogenesis in both familial forms of AF and AF in the general population. We identified a mutation in the cardiac transcription factor NFATC1 in a high-risk Utah pedigree defined by young onset AF. NFATC1 is a Ca2+- dependent transcription factor postulated to play a role in cardiac development, but up until now has never been associated with cardiac excitability. Our exciting preliminary data identify a novel role for NFATC1 in modulating atrial excitability, in that nfatc1 null zebrafish develop spontaneous atrial tachyarrhythmia and juvenile sudden death. Aim 1 seeks to define the biochemical, molecular and electrophysiological basis of the mutant NFATC1 dysfunction, using the HL-1 atrial cell line, a human cell culture model (patient-specific and genome-edited induced pluripotent stem cell derived cardiomyocytes, iPSC-CMs), and a whole animal model (transgenic zebrafish). Aim 2 characterizes the role of NFATC1 in modulating atrial excitability by exploring the molecular and electrophysiological basis for the atrial tachyarrhythmia in nfatc1 null zebrafish. Aim 3 will define NFATC1- controlled transcriptional networks and gene pathways that regulate cardiac excitability, using single-cell RNA- Seq, ChIP-Seq and ChIP-Mass Spectroscopy in human atrium and NFATC1 null and WT iPSC-CMs. Our proposal leverages human genetics, genome-editing techniques, state-of-the-art Next-Gen sequencing modalities and bioinformatics, and electrophysiology to comprehensively characterize the novel role of NFATC1 in cardiac excitability and AF pathogenesis. An understanding of a key transcriptional network that regulates ion channel gene expression and atrial excitability will provide a broader, and more comprehensive understanding of arrhythmia susceptibility and lay the foundation for novel AF therapies.